The immediate aim of this research program is the development of cell biological assays at relatively quantitative levels of resolution for application to cell suspensions and dissociated monolayers of cultured nerve, endocrine and immune cells. The techniques in development include dual-laser fluorescence-activated cell analysis and sorting (FACS), light- and electron-microscopic study of cultured elements using both cytoplasmic and surface-reactive immunologic probes and computer-assisted, quantitative analysis of fluorescence signals from living cells in monolayer culture. Principal observations this year include: 1) a significant and seemingly paradoxical shift in the FACS light-scatter histogram of live cells during embryonic development of the mouse spinal cord but not in histograms of other CNS regions; 2) routine retrograde labeling and FACS isolation of motoneurons and sensory cells followed by long-term culture and morphological and electrophysiological characterization; 3) immunostaining of live prolactin-secreting pituitary cells using anti-prolactin antibody followed by FACS isolation and culture; 4) surface-immunostaining of live embryonic mesencephalic cells followed by FACS analysis, isolation, culture and immunocytochemical characterization with enrichment for catecholaminergic neurons; 5) immunostaining and morphological characterization of dopamine-containing neurons in the spinal cord both in vitro and in vivo; 6) computer-assisted quantitative analysis of fluorescence signals in single nerve, endocrine and immune cells; 7) FACS analysis of effector lymphocytes and their conjugation with tumor target cells. The techniques and protocols used in these projects represent complementary ways of assaying functionally important properties in different cellular phenotypes in a quantitative manner at the single-cell level of experimental resolution.